Microbial fertilizer for soil remediation containing Bacillus subtilis

Abstract

The invention discloses a microbial fertilizer for soil remediation containing bacillus subtilis, belongs to the technical field of fertilizers, and comprises the following raw materials in parts by weight: 30-40 parts of modified biochar, 20-30 parts of bacillus subtilis fermentation liquid, 2-5 parts of trehalose, 20-25 parts of bentonite, 5-10 parts of oxygenator, 3-5 parts of gypsum, and 10 parts of water; the bacillus subtilis provided by the invention is a dominant strain screened out by streaking separation through high-throughput sequencing in a Nanling lotus pond reclamation test field, can quickly repair reclaimed soil, can improve crop resistance and increase crop yield, and the microbial fertilizer prepared by using the bacillus subtilis as a living organism raw material can effectively improve the nutrient content of the reclaimed soil, has a significant effect on improving the organic matter and effective nutrient content of the reclaimed soil, can also increase the crop biomass, and reduce the heavy metal content in the crop.

Description

Technical Field

[0001] The invention belongs to the technical field of fertilizers, and in particular relates to a microbial fertilizer for soil remediation containing Bacillus subtilis. Background Art

[0002] With the large-scale mining in mining areas, the problems of land waste and ecological environment damage caused by mining are becoming increasingly serious. In this regard, reclamation work has received widespread attention. The reclaimed soil has been degraded due to the destruction of the original soil sequence and compaction by large machinery during the reclamation process. The soil is relatively poor and cannot be cultivated directly. Fertilizers need to be added for improvement.

[0003] At present, the fertilizers used for land reclamation in coal mining subsidence areas are generally inorganic fertilizers or organic-inorganic compound fertilizers. The main problems are the lack of living organisms, lack of microorganisms in the soil, slow material energy conversion, slow improvement and recovery of the soil's material properties, biological properties and other soil ecosystems, long reclamation cycle, and excessive use will lead to soil compaction, weakening of soil fertility, eutrophication of water bodies and reduced quality of agricultural products. Microbial fertilizers are safe, efficient and environmentally friendly living microbial products that can effectively increase the content of nitrogen, phosphorus, potassium and other elements in the soil. However, the existing microbial fertilizers have no obvious effect on the improvement of soil reclamation in mining areas. Summary of the invention

[0004] The purpose of the present invention is to provide a microbial fertilizer for soil remediation containing Bacillus subtilis, so as to solve the problem that the existing microbial fertilizer has an insignificant improvement effect on the reclaimed soil in mining areas.

[0005] The purpose of the present invention can be achieved through the following technical solutions:

[0006] A microbial fertilizer for soil remediation containing Bacillus subtilis, comprising the following raw materials in parts by weight:

[0007] 30-40 parts of modified biochar, 20-30 parts of Bacillus subtilis fermentation liquid, 2-5 parts of trehalose, 20-25 parts of bentonite, 5-10 parts of oxygenator, 3-5 parts of gypsum and 10 parts of water.

[0008] Furthermore, the method for preparing the Bacillus subtilis fermentation broth comprises the following steps:

[0009] Bacillus subtilis was inoculated into liquid culture medium and cultured at 37-40°C for 52-56h to obtain a Bacillus subtilis content of 3.5×10 8 cfu / mL-6.5×10 8 cfu / mL of fermentation broth.

[0010] Furthermore, the liquid culture medium uses water as a solvent and includes the following components in concentrations: 8-12 g / L starch, 2-6 g / L calcium chloride, 2-6 g / L yeast extract, 2-6 g / L tryptone, 1.5-3.5 g / L ammonium chloride, and pH = 6.5-7.5.

[0011] Furthermore, the Bacillus subtilis is Bacillus subtilis YJ001, the preservation location is China Center for Type Culture Collection, the preservation time is November 22, 2021, the preservation number is CCTCC NO: M 20211461, and its 16SrDNA sequence is shown in SEQ ID NO.1.

[0012] Furthermore, the modified biochar is γ-polyglutamic acid modified biochar.

[0013] Furthermore, the oxygen-increasing agent is polydopamine-embedded nano-calcium peroxide.

[0014] Furthermore, the method for preparing modified biochar comprises the following steps:

[0015] The biochar was immersed in sodium hydroxide solution and stirred for 1 hour, then shaken for 12 hours, filtered and dried to obtain pretreated biochar; γ-polyglutamic acid was added to deionized water, stirred at 20-40°C for 10 minutes, then added to the pretreated biochar, stirred at 50°C for 10 minutes, glutaraldehyde was added, and continued to stir for 90 minutes, filtered, the filter cake was washed with deionized water and dried to obtain modified biochar.

[0016] Biochar has high carbon content, good stability, rich surface functional groups, developed pores, high specific surface area and cation exchange capacity, can provide nutrition and effective shelter for the survival of microorganisms, and has the potential to improve soil fertility and fix carbon. It is widely used as a bacterial carrier. However, the adsorption capacity of existing biochar on bacteria is limited, which is not conducive to the preparation of efficient microbial fertilizers. In this regard, the present invention uses γ-polyglutamic acid to activate and modify biochar, and introduces -COOH, C=O and -NH on the surface of activated carbon. 2 Functional groups increase the amount of bacteria absorbed. In addition, γ-polyglutamic acid has good water retention and adhesion, which can improve the microenvironment in which the bacteria are located, reduce the interference and damage of the external environment to Bacillus subtilis, and contribute to the survival of the bacteria.

[0017] Furthermore, the usage ratio of γ-polyglutamic acid, deionized water, pretreated biochar and glutaraldehyde is 0.15-0.3 g:20 mL:1 g:4 mL.

[0018] Furthermore, the concentration of the sodium hydroxide solution during the preparation of the pretreatment biochar was 1 mol / L.

[0019] Furthermore, the molecular weight of γ-polyglutamic acid is 50 kDa.

[0020] Furthermore, the biochar was corn straw biochar prepared by pyrolysis at 400°C and was purchased from Henan Boton Biotechnology Co., Ltd.

[0021] Furthermore, the preparation method of the oxygenator comprises the following steps:

[0022] Dissolve calcium chloride in deionized water, add ammonia water dropwise to adjust the pH value to 10±0.5 to obtain a mixed solution a, add hydrogen peroxide solution to the mixed solution a, stir evenly, add sodium hydroxide to adjust the pH value to 11 to obtain a mixed solution b, add dopamine hydrochloride to the mixed solution b, stir evenly, adjust the pH value to 8.5 with sodium hydroxide, continue stirring for 2 hours, centrifuge, wash the precipitate with anhydrous ethanol for 3 times, and then vacuum dry at 60°C to constant weight to obtain an oxygenator.

[0023] Calcium peroxide has good oxidizing properties and can be used as a source of hydrogen peroxide. It also has the function of releasing oxygen. The reaction equation with water is as follows:

[0024] CaO 2 +2H 2 O=Ca(OH) 2 +H 2 O 2 ;

[0025] CaO 2 +2H 2 O=2Ca(OH) 2 +O 2 ↑;

[0026] Based on this, the present invention uses calcium peroxide as an oxygen source and polydopamine as an embedding agent to prepare polydopamine-embedded nano calcium peroxide, i.e., the oxygenator. By embedding treatment, the oxygenator can slowly release O when in contact with water. 2 and H 2 O 2 .

[0027] Furthermore, the dosage ratio of calcium chloride, deionized water, hydrogen peroxide solution and dopamine hydrochloride is 2g:20-30mL:10mL:0.8-1.1g, and the mass fraction of hydrogen peroxide solution is 30%.

[0028] Furthermore, the mass fraction of ammonia water is 25-28%.

[0029] The method for preparing the microbial fertilizer for soil remediation containing Bacillus subtilis comprises the following steps:

[0030] The modified organism, Bacillus subtilis fermentation liquid, trehalose, bentonite, oxygenator, gypsum and water are added into a mixer and mixed evenly, and then transferred into a disc granulator for granulation to obtain fertilizer granules with a diameter of 1-3.5 mm, and then dried and shaped.

[0031] Furthermore, the drying and setting temperature is set to 30°C-50°C, and the drying and setting time is 2-8h.

[0032] Beneficial effects of the present invention:

[0033] 1. The Bacillus subtilis provided by the present invention is a dominant strain screened out from the Nanling Lotus Pond reclamation test field through high-throughput sequencing and then streaking separation. It can quickly repair the reclaimed soil, improve crop resistance and increase crop yield. The microbial fertilizer prepared with it as a living organism raw material can effectively increase the nutrient content of the reclaimed soil, has a significant effect on improving the organic matter and effective nutrient content of the reclaimed soil, and can also increase the crop biomass and reduce the heavy metal content in the crop.

[0034] 2. The present invention uses γ-polyglutamic acid-modified biochar as a carrier for loading Bacillus subtilis. Compared with unmodified biochar, it has the characteristics of large bacterial quantity and high survival rate of bacteria, can maintain the effective content of Bacillus subtilis in microbial fertilizer, better improve the properties of reclaimed soil, and increase crop yield. In addition, γ-polyglutamic acid has a strong binding ability for heavy metal ions in the soil, passivates the heavy metal ions in the reclaimed soil, and reduces the absorption of heavy metal ions by crops.

[0035] 3. The present invention adds an oxygenator to the microbial fertilizer, which can slowly release oxygen in the soil, significantly improves the oxygen content in the soil, provides a more suitable living environment for beneficial microorganisms such as Bacillus subtilis, promotes the rapid reproduction and activity enhancement of beneficial bacteria such as Bacillus subtilis, thereby improving the effect of the microbial fertilizer, and can also improve the soil structure, increase the soil porosity, indirectly improve the air permeability and water permeability of the soil, help root respiration and water penetration, and create a healthier soil microenvironment for plant growth. The surface of the oxygenator also contains amino and catechol structures, which can chelate heavy metal ions, passivate the heavy metal ions in the soil, and further reduce the absorption of heavy metal ions by crops. DETAILED DESCRIPTION

[0036] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0037] In the present application, the molecular weight of γ-polyglutamic acid is 50 kDa; the biochar is corn straw biochar prepared by pyrolysis at 400° C., which was purchased from Henan Boton Biotechnology Co., Ltd.

[0038] Preparation Example 1

[0039] The method for preparing modified biochar comprises the following steps:

[0040] 10 g of biochar was soaked and stirred in 100 mL of 1 mol / L sodium hydroxide solution for 1 h, then shaken for 12 h, filtered and dried to obtain pretreated biochar; 0.15 g of γ-polyglutamic acid was added to 20 mL of deionized water, stirred at 20 ° C for 10 min, then 1 g of pretreated biochar was added, stirred at 50 ° C for 10 min, 4 mL of glutaraldehyde was added, and stirring continued for 90 min, filtered, the filter cake was washed with deionized water and dried to obtain modified biochar.

[0041] Preparation Example 2

[0042] The method for preparing modified biochar comprises the following steps:

[0043] 10 g of biochar was soaked and stirred in 100 mL of 1 mol / L sodium hydroxide solution for 1 h, then shaken for 12 h, filtered and dried to obtain pretreated biochar; 0.3 g of γ-polyglutamic acid was added to 20 mL of deionized water, stirred at 40 ° C for 10 min, then 1 g of pretreated biochar was added, stirred at 50 ° C for 10 min, 4 mL of glutaraldehyde was added, and stirring was continued for 90 min, filtered, the filter cake was washed with deionized water and dried to obtain modified biochar.

[0044] Comparative Example 1

[0045] This control example is biochar.

[0046] Comparative Example 2

[0047] The preparation method of the modified biochar is different from that of Preparation Example 1 in that 0.15 g of γ-polyglutamic acid is changed to 0.1 g of γ-polyglutamic acid.

[0048] Comparative Example 3

[0049] The preparation method of the modified biochar is different from that of Preparation Example 2 in that 0.3 g of γ-polyglutamic acid is changed to 0.35 g of γ-polyglutamic acid.

[0050] Example 1

[0051] 1. Isolation of bacterial strains

[0052] Weigh 10g of soil sample from the Hetang reclamation test field in Nanling, Anhui Province, add it to a conical flask containing 90mL of sterile saline and glass beads, shake it on a shaker at 180rpm for 30min, and then place it in an 80℃ water bath for 20min to inhibit the growth of bacteria. After ten-fold gradient dilution, take 100μL of each sample from the dilution and evenly spread it on the LB plate medium (nutrient broth solid plate). The coated plate is placed in a 37℃ constant temperature incubator for 12-24h, and a single suspected Bacillus subtilis colony formed on the LB plate is picked.

[0053] The single colony was purified by streaking multiple times on a fresh LB plate by the streaking method until there was no interference from other bacteria under a microscope. A target strain with morphology and characteristics consistent with those of Bacillus subtilis was successfully obtained and named YJ001.

[0054] 2. Identification of bacterial strains

[0055] (I) Identification based on morphological and physiological and biochemical characteristics

[0056] According to the standard methods in Bergey's Manual of Bacterial Identification (8th Edition) and Manual of Systematic Identification of Common Bacteria, the morphological characteristics, culture characteristics, and physiological and biochemical characteristics of strain YJ001 were systematically analyzed. The results are as follows:

[0057] Morphology of strains:

[0058] After culturing on LB medium at 37°C for 24 hours, the colony morphology is obvious. The colony is light yellow, nearly round, dry and dull, with obvious wrinkled protrusions and irregular edges. Under the microscope, the bacteria are straight rod-shaped, Gram-positive, and form oval meso-spores.

[0059] Physiological and biochemical characteristics:

[0060] Gelatin liquefaction: positive (+); catalase activity: positive (+); VP reaction: positive (+); indole formation: negative (-); citrate utilization: negative (-); glucose fermentation and gas production: positive (+); xylose gas production: positive (+), but no acid production; mannitol gas production: positive (+), but no acid production; arabinose acid production: positive (+); starch hydrolysis: positive (+); casein hydrolysis: positive (+); tyrosine hydrolysis: negative (-); nitrate reduction: positive (+); Salt tolerance: can grow well under 2%, 5%, and 7% NaCl conditions.

[0061] (II) 16S rDNA test

[0062] Single colonies on the plate were picked and identified by 16S rRNA colony PCR. The purified PCR products were sent to Guangdong Meige Gene Technology Co., Ltd. for gene sequencing, and the sequence measured was SEQ ID NO. 1. According to the sequence homology comparison of the GenBank database, the homology between strain YJ0011 and Bacillus subtilis strain GX S-11 (GenBank accession number KU904283.1) reached 99%, and the strain was determined to be Bacillus subtilis.

[0063] Based on the above results, the strain was named Bacillus subtilis YJ001 and deposited in the China Center for Type Culture Collection on November 22, 2021, with the preservation number CCTCC NO: M 20211461. The preservation address is No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province.

[0064] Example 2

[0065] A microbial fertilizer for soil remediation containing Bacillus subtilis, comprising the following raw materials in parts by weight:

[0066] 30 parts of the modified biochar of Preparation Example 1, 20 parts of Bacillus subtilis fermentation liquid, 2 parts of trehalose, 20 parts of bentonite, 5 parts of oxygenator, 3 parts of gypsum, and 10 parts of water.

[0067] The method for preparing Bacillus subtilis fermentation broth comprises the following steps:

[0068] Bacillus subtilis was inoculated into liquid culture medium and cultured at 37°C for 56 h to obtain a Bacillus content of 3.5×10 8 cfu / mL of fermentation broth.

[0069] The liquid culture medium uses water as solvent and includes components in the following concentrations: 8 g / L starch, 2 g / L calcium chloride, 2 g / L yeast extract, 2 g / L tryptone, 1.5 g / L ammonium chloride, and pH=6.5.

[0070] The preparation method of the oxygenator comprises the following steps:

[0071] Dissolve 2 g of calcium chloride in 20 mL of deionized water, add 10 mL of 25 wt % ammonia water to adjust the pH value to 10 to obtain a mixed solution a, add 10 mL of 30 wt % hydrogen peroxide solution to the mixed solution a, stir evenly, add sodium hydroxide to adjust the pH value to 11 to obtain a mixed solution b, add 0.8 g of dopamine hydrochloride to the mixed solution b, stir evenly, adjust the pH value to 8.5 with sodium hydroxide, continue stirring for 2 h, centrifuge, wash the precipitate with anhydrous ethanol 3 times, and then vacuum dry at 60 ° C to constant weight to obtain an oxygenator.

[0072] The method for preparing the microbial fertilizer for soil remediation containing Bacillus subtilis comprises the following steps:

[0073] Add the modified organism, Bacillus subtilis fermentation liquid, trehalose, bentonite, oxygenator, gypsum and water into a mixer and mix evenly, transfer to a disc granulator for granulation to obtain fertilizer particles with a diameter of 1-3.5 mm, and then dry at 30° C. for 8 hours.

[0074] Example 3

[0075] A microbial fertilizer for soil remediation containing Bacillus subtilis, comprising the following raw materials in parts by weight:

[0076] 35 parts of the modified biochar of Preparation Example 1, 25 parts of Bacillus subtilis fermentation liquid, 3 parts of trehalose, 22 parts of bentonite, 8 parts of oxygenator, 4 parts of gypsum, and 10 parts of water.

[0077] The method for preparing Bacillus subtilis fermentation broth comprises the following steps:

[0078] Bacillus subtilis was inoculated into liquid culture medium and cultured at 38°C for 54 h to obtain a Bacillus content of 5.0×10 8 cfu / mL of fermentation broth.

[0079] The liquid culture medium uses water as solvent and includes components in the following concentrations: 10 g / L starch, 4 g / L calcium chloride, 4 g / L yeast extract, 4 g / L tryptone, 3 g / L ammonium chloride, and pH=7.0.

[0080] The preparation method of the oxygenator comprises the following steps:

[0081] Dissolve 2 g of calcium chloride in 25 mL of deionized water, add 10 mL of 27 wt % ammonia water to adjust the pH value to 10 to obtain a mixed solution a, add 10 mL of 30 wt % hydrogen peroxide solution to the mixed solution a, stir evenly, add sodium hydroxide to adjust the pH value to 11 to obtain a mixed solution b, add 1 g of dopamine hydrochloride to the mixed solution b, stir evenly, adjust the pH value to 8.5 with sodium hydroxide, continue stirring for 2 h, centrifuge, wash the precipitate with anhydrous ethanol 3 times, and then vacuum dry at 60 ° C to constant weight to obtain an oxygenator.

[0082] The method for preparing the microbial fertilizer for soil remediation containing Bacillus subtilis comprises the following steps:

[0083] Add the modified organism, Bacillus subtilis fermentation liquid, trehalose, bentonite, oxygenator, gypsum and water into a mixer and mix evenly, transfer to a disc granulator for granulation to obtain fertilizer particles with a diameter of 1-3.5 mm, and then dry at 40° C. for 6 hours.

[0084] Example 4

[0085] A microbial fertilizer for soil remediation containing Bacillus subtilis, comprising the following raw materials in parts by weight:

[0086] 40 parts of the modified biochar of Preparation Example 2, 30 parts of Bacillus subtilis fermentation liquid, 5 parts of trehalose, 25 parts of bentonite, 10 parts of oxygenator, 5 parts of gypsum, and 10 parts of water.

[0087] The method for preparing Bacillus subtilis fermentation broth comprises the following steps:

[0088] Bacillus subtilis was inoculated into liquid culture medium and cultured at 40°C for 56 h to obtain a Bacillus content of 6.5×10 8 cfu / mL of fermentation broth.

[0089] The liquid culture medium uses water as a solvent and includes components with the following concentrations: 12 g / L starch, 6 g / L calcium chloride, 6 g / L yeast extract, 6 g / L tryptone, 3.5 g / L ammonium chloride, and pH=7.5.

[0090] The preparation method of the oxygenator comprises the following steps:

[0091] Dissolve 2 g of calcium chloride in 30 mL of deionized water, add 10 mL of 28 wt % ammonia water to adjust the pH value to 10.5 to obtain a mixed solution a, add 10 mL of 30 wt % hydrogen peroxide solution to the mixed solution a, stir evenly, add sodium hydroxide to adjust the pH value to 11 to obtain a mixed solution b, add 1.1 g of dopamine hydrochloride to the mixed solution b, stir evenly, adjust the pH value to 8.5 with sodium hydroxide, continue stirring for 2 h, centrifuge, wash the precipitate with anhydrous ethanol 3 times, and then vacuum dry at 60 ° C to constant weight to obtain an oxygenator.

[0092] The method for preparing the microbial fertilizer for soil remediation containing Bacillus subtilis comprises the following steps:

[0093] Add the modified organism, Bacillus subtilis fermentation liquid, trehalose, bentonite, oxygenator, gypsum and water into a mixer and mix evenly, transfer to a disc granulator for granulation to obtain fertilizer particles with a diameter of 1-3.5 mm, and then dry at 50° C. for 2 hours.

[0094] Comparative Example 1

[0095] A microbial fertilizer for soil remediation containing Bacillus subtilis. Compared with Example 2, the modified biochar in Example 2 is replaced by the substance in Control Example 1, and the remaining raw materials and preparation process are the same as in Example 2.

[0096] Comparative Example 2

[0097] A microbial fertilizer for soil remediation containing Bacillus subtilis. Compared with Example 2, the modified biochar in Example 2 is replaced by the product prepared in Control Example 2, and the remaining raw materials and preparation process are the same as in Example 2.

[0098] Comparative Example 3

[0099] A microbial fertilizer for soil remediation containing Bacillus subtilis. Compared with Example 4, the modified biochar in Example 4 is replaced by the product prepared in Control Example 3, and the remaining raw materials and preparation process are the same as Example 4.

[0100] Comparative Example 4

[0101] A microbial fertilizer for soil remediation containing Bacillus subtilis. Compared with Example 2, the oxygenator in Example 2 is replaced by calcium peroxide, and the remaining raw materials and preparation process are the same as Example 2.

[0102] Comparative Example 5

[0103] A microbial fertilizer for soil remediation containing Bacillus subtilis. Compared with Example 2, the oxygenator in Example 2 is removed, and the remaining raw materials and preparation process are the same as Example 2.

[0104] (I) The microbial fertilizers prepared in Examples 2-4 and Comparative Examples 1-3 were placed in sealed bags, and the effective viable bacterial counts were measured at 1 day, 7 days, 30 days, and 180 days, respectively. The effective viable bacterial counts were measured in accordance with the effective viable bacterial count measurement method specified in the microbial fertilizer product inspection procedures NY / T2321-2013. The test results are shown in Table 1:

[0105] Table 1

[0106]

[0107] It can be seen from the data recorded in Table 1 that the effective live bacteria counts at 1d, 7d, and 30d in the microbial fertilizers of Examples 2-4 are all greater than those of Comparative Examples 1-3, indicating that the microbial fertilizer of the present invention not only has a high effective live bacteria count but also has good storage properties. Specifically, it can be seen from the test results of Example 2 and Comparative Example 1 that the microbial fertilizer prepared using modified biochar has a high effective live bacteria count and good storage properties compared to biochar. It can be seen from the test results of Example 2, Comparative Example 2 and Example 4, Comparative Example 3 that a too low or too high proportion of γ-polyglutamic acid in the preparation process of modified biochar will have an adverse effect, mainly affecting the porosity of the biochar, causing the degradation of the living environment of the bacterial agent, and ultimately leading to a low effective live bacteria count in the microbial fertilizer.

[0108] (II) Effect of microbial fertilizers prepared in Examples 2-4 and Comparative Examples 1-5 on soil improvement in coal mining subsidence areas

[0109] (1) Potted plant test

[0110] The test soil samples were collected from the soil around the coal mining subsidence area in Bengbu City, Anhui Province. The test vegetables were dwarf yellow. A total of 8 treatments were set up in the test. The microbial fertilizers prepared by Examples 2-4 and Comparative Examples 1-5 were applied respectively, and the application amount was 4g / kg soil (4‰). The potting container is a 150mm×125mm (upper diameter×height) plastic pot, and each pot contains 1.0kg of air-dried soil. The microbial fertilizers of each group were evenly mixed with the soil according to the application amount of the above treatment, and then potted, and watered thoroughly before sowing. The seeds that have been germinated and whitened were sown in pots, 6 per pot. Keep the soil moist from sowing to emergence, water once 2-3 days after emergence, and maintain 40% of the maximum field water holding capacity. When 2-3 true leaves grow, thinning is carried out, the seedling spacing is 3-4cm, 2 plants are left in each pot, and water is timely watered once after thinning. Harvest after 40 days, and collect soil and plant samples. Basic soil physical and chemical properties such as soil organic carbon, available nitrogen, phosphorus and potassium were determined. The results are shown in Table 2. Vegetable biomass and Cd in vegetable leaves were determined. 2+ The results are shown in Table 3.

[0111] Table 2

[0112]

[0113] From the data recorded in Table 2, it can be seen that compared with the reclaimed soil improved by microbial soil in Comparative Examples 1-5, the physical and chemical properties of the soil in Examples 2-4 are more significantly improved, and the soil organic matter content and the content of effective nutrients are significantly increased.

[0114] Table 3

[0115]

[0116] From the data recorded in Table 3, it can be seen that compared with the vegetables planted in the reclaimed soil improved by microbial soil in Comparative Examples 1-5, the vegetables planted in the reclaimed soil improved by microbial soil in Examples 2-4 have larger fresh weight and dry weight, and the Cd content in the leaves is 2+ Lower content.

[0117] It should be noted that, in this article, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprise" or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device.

[0118] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A microbial fertilizer for soil remediation containing Bacillus subtilis, characterized in that: It includes the following raw materials in parts by weight: 30-40 parts of modified biochar, 20-30 parts of Bacillus subtilis fermentation liquid, 2-5 parts of trehalose, 20-25 parts of bentonite, 5-10 parts of oxygenator, 3-5 parts of gypsum, and 10 parts of water; The modified biochar is γ-polyglutamic acid modified biochar; The oxygenator is polydopamine-embedded nano calcium peroxide; The Bacillus subtilis in the Bacillus subtilis fermentation broth is classified and named Bacillus subtilis YJ001, which is deposited in the China Center for Type Culture Collection with a deposit number of CCTCC NO: M 20211461, and its 16S rDNA sequence is shown in SEQ ID NO.1; The content of Bacillus subtilis in the Bacillus subtilis fermentation broth was 3.5×10 8 cfu / mL-6.5×10 8 cfu / mL; The method for preparing modified biochar comprises the following steps: The biochar was immersed in sodium hydroxide solution and stirred for 1 hour, then shaken for 12 hours, filtered and dried to obtain pretreated biochar; γ-polyglutamic acid was added to deionized water, stirred at 20-40°C for 10 minutes, and then the pretreated biochar was added, stirred at 50°C for 10 minutes, glutaraldehyde was added, and continued to stir for 90 minutes, filtered, and the filter cake was washed and dried to obtain modified biochar; The dosage ratio of γ-polyglutamic acid, deionized water, pretreated biochar, and glutaraldehyde was 0.15-0.3 g: 20 mL: 1 g: 4 mL; The molecular weight of γ-polyglutamic acid is 50 kDa.

2. The microbial fertilizer for soil remediation containing Bacillus subtilis according to claim 1, characterized in that: The preparation method of the oxygenator comprises the following steps: Dissolve calcium chloride in deionized water, add ammonia water dropwise to adjust the pH value to 10±0.5 to obtain a mixed solution a, add hydrogen peroxide solution to the mixed solution a, stir evenly, add sodium hydroxide to adjust the pH value to 11 to obtain a mixed solution b, add dopamine hydrochloride to the mixed solution b, stir evenly, adjust the pH value to 8.5 with sodium hydroxide, continue stirring for 2 hours, centrifuge, wash the precipitate and dry it to obtain an oxygenator.

3. The microbial fertilizer for soil remediation containing Bacillus subtilis according to claim 2, characterized in that: The dosage ratio of calcium chloride, deionized water, hydrogen peroxide solution and dopamine hydrochloride is 2g:20-30mL:10mL:0.8-1.1g, and the mass fraction of hydrogen peroxide solution is 30%.

Images